This invention relates to sensors for performing electrochemical assays for the accurate determination of one or more enzymes or their substrates in complex liquid mixtures. The invention is of particular utility for biomedical applications in whole blood.
The measurement of dissolved analytes in whole blood is subject to interference by red cells (hematocrit) due to the volume exclusion of the non-liquid fraction of the cells. In addition, where a rate reaction based upon the turnover of the analytes by a specific enzyme is utilized to measure analyte concentration, measurement is subject to the restriction of the diffusional pathways for the analyte and enzyme by the red cell fraction. This interference tends to cause an artificially high response rate for low hematocrit levels, and, conversely, an artificially low response rate for high hematocrit levels.
A common method for overcoming this interference is to filter out or otherwise separate off the red cells and measure the concentration of the analyte in plasma, or to pretreat the whole blood sample to reduce or eliminate the red cell volume (e.g. by lysis of the cells). However, the incorporation of a sample pre-treatment step or the requirement to separate off the red cell fraction in a whole blood sample does not give rise to measurement devices which are simple and convenient to use outside of the analytical laboratory.
Another problem encountered with some electrode strips used with electrochemical sensors is that application of an insufficient amount of sample to the electrodes tends to produce an artificially low response due to the failure to completely cover the working electrode with sample. Moreover, even if a sufficient amount of sample is initially applied to the strip, if the strip is not held in a horizontal position the sample can flow off of the strip, and/or the electrode area may not be wetted by the sample due to the surface tension of the sample.
U.S. patent applications Ser. Nos. 08/281,131 and 08/281,237 and European Patents Nos. 127,958 and 351,891 describe the construction of sensors comprising a conductive electrode coated with a mixture, or layers, of a catalytically active enzyme and a mediator compound. When such a coated electrode is contacted with a liquid sample containing a species for which the enzyme exerts a catalytic effect, the mediator compound transfers electrons from the reaction and the resulting current can be used to give a readout signal relating to the concentration of the said substrate. Alternatively, a substrate can be used in the mixture instead of an enzyme when it is desired to measure an enzyme which exerts a catalytic effect on the substrate.
Such sensors are used in the chemical industry, e.g., to analyze complex mixtures, for example in the food industry and in biochemical engineering, and are also valuable in biological investigation in human or animal medicine. The sensors may be used as invasive probes, i.e., one that can be inserted into the body and make contact with a body fluid such as whole blood or subcutaneous fluid. Alternatively the electrode system may be used as part of an external test upon a withdrawn sample (e.g., blood withdrawn by a syringe) or upon an expressed sample (e.g., blood obtained using a needle pricking device).